Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The urokinase-type plasminogen activator receptor (uPAR) plays an important role on the cell surface in mediating extracellular degradative processes and formation of active TGF-beta, and in nonproteolytic events such as cell adhesion, migration, and transmembrane signaling. We have searched for mechanisms that determine the cellular location of uPAR and may participate in its disposal. When using purified receptor preparations, we find that uPAR binds to the cation-independent, mannose 6-phosphate/insulin-like growth factor-II (IGF-II) receptor (CIMPR) with an affinity in the low micromolar range, but not to the 46-kD, cation-dependent, mannose 6-phosphate receptor (CDMPR). The binding is not perturbed by uPA and appears to involve domains DII + DIII of the uPAR protein moiety, but not the glycosylphosphatidylinositol anchor. The binding occurs at site(s) on the CIMPR different from those engaged in binding of mannose 6-phosphate epitopes or IGF-II. To evaluate the significance of the binding, immunofluorescence and immunoelectron microscopy studies were performed in transfected cells, and the results show that wild-type CIMPR, but not CIMPR lacking an intact sorting signal, modulates the subcellular distribution of uPAR and is capable of directing it to lysosomes. We conclude that a site within CIMPR, distinct from its previously known ligand binding sites, binds uPAR and modulates its subcellular distribution.
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PMID:Mannose 6-phosphate/insulin-like growth factor-II receptor targets the urokinase receptor to lysosomes via a novel binding interaction. 956 79

During acute and chronic inflammatory lung diseases, the normal fibrinolytic activity in the alveolar space is inhibited by increased levels of plasminogen activator inhibitor 1 (PAI-1). Transgenic mice having increased fibrinolytic activity due to genetic deficiency of PAI-1 develop less fibrosis after bleomycin-induced lung inflammation. These observations led us to hypothesize that pulmonary fibrosis could be limited through enhancement of alveolar fibrinolytic activity by adenovirus-mediated transfer of the urokinase-type plasminogen activator (uPA) gene to the lung. To investigate this hypothesis, 0.075 U of bleomycin was introduced intratracheally into mice. Twenty-one days later, the mice were treated intratracheally with phosphate-buffered saline (PBS), a control adenovirus, or adenoviruses containing murine or human uPA cDNAs. On day 28, the mice were sacrificed, and lung fibrosis was quantitated by measuring hydroxyproline content. As expected, bleomycin caused a doubling in lung hydroxyproline to 345.6+/-28.2 microg/lung (SEM) compared with mice receiving PBS (170.2+/-4.0 microg/lung). Treatment of the bleomycin-injured mice with the control adenovirus on day 21 had no impact on lung fibrosis (338.4+/-17.2 microg/lung). Importantly, the human uPA adenovirus significantly reduced (p<0.05) lung hydroxyproline (281.2+/-22.8 microg/lung), thus attenuating by 38% the bleomycin-induced increase in lung collagen. The improvement in bleomycin-induced lung fibrosis resulting from treatment with the human uPA adenovirus further supports the importance of the fibrinolytic system during inflammatory lung injury and repair.
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PMID:Treatment of bleomycin-induced pulmonary fibrosis by transfer of urokinase-type plasminogen activator genes. 1051 51

The urokinase receptor (uPAR) is a glycolipid anchored cell surface glycoprotein that plays a central role in extracellular proteolysis during tissue remodeling processes and cancer invasion. By intramuscular (i.m.) injection of rabbits with plasmid DNA coding for a carboxy-terminally truncated secreted form of the murine uPAR (muPAR), specific anti-sera with a titer of 64,000, as measured by ELISA, have been obtained. Rabbits received a total of 10 monthly injections of 1 mg DNA in phosphate-buffered saline. The antibody titer peaked between the 5th and 7th injection and slowly declined after the 8th injection. After the final immunization the immune response persisted for at least 6 months without further injections. The antibodies generated by DNA immunization were useful for immunohistochemistry and immunoblotting, recognizing the antigen both in its native and in its reduced and alkylated form. Using the antibodies in immunoblotting muPAR was identified in lysates of peritoneal macrophages, spleen and lung tissue. Both the intact and cleaved form of muPAR were identified in lysates of a murine monocyte cell line P388D.1. No cross-reaction with human uPAR was observed. In immunohistochemical analysis of normal mouse lung tissue uPAR immunoreactivity was located in the alveoli and pulmonary vessels, whereas the bronchial epithelium was negative. These results demonstrate that DNA immunization of rabbits using i.m. injection is a very effective and easy method to raise polyclonal antibodies which can be used for characterization and localization of muPAR in mouse tissue.
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PMID:Generation of high-affinity rabbit polyclonal antibodies to the murine urokinase receptor using DNA immunization. 1066 75

Hemostasis, the system of generation and degradation of thrombi, consists of coagulation and fibrinolysis. Whereas global assays to study coagulation have existed for many years, there has been no simple, rapid, and economic routine test for the plasmatic fibrinolysis parameters plasminogen activator inhibitor-1, alpha2-antiplasmin, plasminogen, and aprotinin. Here a fast functional global assay for these plasmatic fibrinolytic parameters is presented. However, the present assay is not sensitive to physiological concentrations of prourokinase or tissue-type plasminogen activator. The following assay conditions have been found to be optimal: 50 microL of citrated plasma is incubated with 50 microL of 10 IU urinary-type plasminogen activator (urokinase)/mL, 1.1 mmol/L tranexamic acid, 1% polygelin, 0.1% Triton X-100, phosphate-buffered saline, pH 7.4, for 20 min at 37 degrees C (plasmin generation phase). Then 50 microL of 3 mmol/L HD-Nva-CHA-Lys-pNA, 1.05 mol/L KCl is added, and deltaA (405 nm)/10 min (37 degrees C) is determined, by using a microtiterplate reader (plasmin detection phase). The results are calibrated against pooled normal plasma (100% plasmatic fibrinolytic parameters activity). The intra- and interassay coefficients of variation have been found to be less than 5%. The detection limit (sensitivity) of the functional fibrinolysis assay is 5 % of the normal plasmatic fibrinolysis parameters activity. The normal plasmatic fibrinolysis parameters activity is 100%, sigma = 25%. The plasmatic fibrinolysis parameters activity correlates negatively (r = -0.684) with the plasminogen activator inhibitor-1 activity of patient samples. The plasmatic fibrinolysis parameters assay is a simple, rapid, and economic functional test for several clinical relevant fibrinolysis parameters.
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PMID:A simple screening assay for certain fibrinolysis parameters (FIPA). 1067 10

We examined the tumorigenic and metastatic potentials of three human non-small cell lung cancer (NSCLC) cell lines, PC-14, A549 or Lu-99 cell lines suspended in Matrigel-containing phosphate-buffered saline were orthotopically implanted into the lungs of nude mice. The formation of a solitary tumor nodule in the lung was observed after the implantation of all cell lines. Intrapulmonary implantation of PC-14 or Lu-99 cells resulted in spontaneous distant metastases. In contrast, A549 cells caused multiple intrapulmonary metastases to the right and left lobes of the lung without producing visible lymphatic metastasis. We also investigated the expression of matrix metalloproteinases (MMPs), urokinase-type plasminogen activator (u-PA), u-PA receptor (u-PAR) and c-MET in these cell lines in vitro and in vivo. Reverse transcription polymerase chain reaction (RT-PCR) analysis showed that the expression of MMP-2 and membrane-type 1 MMP (MT1-MMP) was elevated in PC-14 as compared with the other two cell lines. In contrast, stronger expression of c-MET was observed in A549 than in PC-14 or Lu-99. These results indicate that differential patterns of metastasis of lung cancer might be associated with differential expression of metastasis-associated molecules. Our orthotopic implantation models display clinical features resembling those of NSCLC, and may provide a useful basis for lung cancer research.
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PMID:Solitary lung tumors and their spontaneous metastasis in athymic nude mice orthotopically implanted with human non-small cell lung cancer. 1100 66

Previously, we demonstrated that amorphous calcium phosphate (ACP), chemical precursor to apatite, strongly interacted with fibrin and facilitated binding of matrix metalloproteinase (MMP)-9, a type IV collagenase. Plasmin-dependent fibrinolysis resulted in coordinate MMP-9 activation. Here we report on the effect(s) of ACP on fibrin degradation and binding of endogenous plasma proteases. Electrophoresis (8.5% SDS-PAGE) revealed that fibrin formed in the presence of ACP demonstrated characteristic gamma-gamma dimers (90-kDa) and beta-monomers (55-kDa), but resisted spontaneous fibrinolysis (72 h, 37 degrees C) or degradation by plasminogen activators (uPA, tPA). Casein zymography revealed an ACP-dependent decrease in fibrin binding of a low molecular weight (Mw) protease triplet (47-, 43-, 42-kDa) and increased fibrin binding of two high Mw proteases (94- and 84-kDa). The low Mw triplet also possessed gelatinolytic activity, but was not an MMP since 1,10-phenanthroline was ineffective as an inhibitor. Fibrin-binding proteases were inhibited to some degree by the serine protease inhibitor aprotinin. Competition/dissociation experiments with epsilon-aminocaproic acid revealed that the low Mw triplet lacked kringle regions whereas the 94- and 84-kDa proteases were tentatively identified and glu-/lys-plasmin(ogen)s. The triplet may, however, represent one or more kringle deficient mini-plasminogen(s), since electrophoretic mobility and substrate specificity was similar to elastase-generated mini-plasminogen. To explore these findings in a clinically relevant setting, a series of plasma samples was collected from a patient with unstable angina prior to, during, and post coronary artery bypass graft (CABG) surgery. Fibrin formed from plasma collected during and immediately post CABG was associated with increased fibrinolytic capacity and enhanced binding of a) MMP-9, b) the low Mw protease triplet (described above), and c) PA (as putative 110-kDa tPA:PAI-1 complex). The relevance of these findings to pathologic calcification of atherosclerotic plaques is discussed.
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PMID:Interaction of amorphous calcium phosphate with fibrin in vitro causes decreased fibrinolysis and altered protease profiles: implications for atherosclerotic disease. 1182 Apr 59

We investigated the specific role of matrix metalloproteinase (MMP)-9 in allergic asthma using a murine model of allergen-induced airway inflammation and airway hyperresponsiveness in MMP-9(-/-) mice and their corresponding wild-type (WT) littermates. After a single intraperitoneal sensitization to ovalbumin, the mice were exposed daily either to ovalbumin (1%) or phosphate-buffered saline aerosols from days 14 to 21. Significantly less peribronchial mononuclear cell infiltration of the airways and less lymphocytes in the bronchoalveolar lavage fluid were detected in challenged MMP-9(-/-) as compared to WT mice. In contrast, comparable numbers of bronchoalveolar lavage fluid eosinophils were observed in both genotypes. After allergen exposure, the WT mice developed a significant airway hyperresponsiveness to carbachol whereas the MMP-9(-/-) mice failed to do so. Allergen exposure induced an increase of MMP-9-related gelatinolytic activity in WT lung extracts. Quantitative reverse transcriptase-polymerase chain reaction showed increased mRNA levels of MMP-12, MMP-14, and urokinase-type plasminogen activator after allergen exposure in the lung extracts of WT mice but not in MMP-9-deficient mice. In contrast, the expression of tissue inhibitor of metalloproteinases-1 was enhanced after allergen exposure in both groups. We conclude that MMP-9 plays a key role in the development of airway inflammation after allergen exposure.
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PMID:Matrix metalloproteinase-9 deficiency impairs cellular infiltration and bronchial hyperresponsiveness during allergen-induced airway inflammation. 1216 74

Leukocyte migration to sites of inflammation is a multistep process involving transient adhesion to the endothelium followed by cell surface-controlled proteolysis for transmigration through the vessel wall and chemotactic movement within tissues. One of the key players in this machinery appears to be the urokinase-type plasminogen activator (uPA)/uPA receptor system. The role of uPA and its receptor (CD87) in plasminogen (Plg) activation, cell adhesion, and chemotaxis is well established; however, less is known of how these activities are regulated. Here we provide evidence that the mannose 6-phosphate/insulin-like growth factor 2 receptor (CD222) controls CD87-mediated functions. Expression of human CD222 in CD222-/- mouse fibroblasts down-regulated Plg activation, cell adhesion, and chemotaxis induced by the uPA/CD87 system. In addition, we demonstrate that the N-terminal region of CD222, which is similar to the Plg-binding site of streptokinase, plays a crucial role in binding of CD87 and Plg. A peptide derived from this region in CD222 is able to disrupt the physical interaction of CD222 with CD87 and, furthermore, mimics the inhibitory effects of CD222 on CD87 functions. Taken together, our results indicate a novel role for CD222 in regulation of fibrinolysis, cell adhesion, and migration.
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PMID:The N terminus of mannose 6-phosphate/insulin-like growth factor 2 receptor in regulation of fibrinolysis and cell migration. 1218 57

Matriptase is an epithelial-derived, cell surface serine protease. This protease activates hepatocyte growth factor (HGF) and urokinase plasminogen activator (uPA), two proteins thought to be involved in the growth and motility of cancer cells, particularly carcinomas, and in the vascularization of tumors. Thus, matriptase may play an important role in the progression of carcinomas, such as breast cancer. We examined the regulation of activation of matriptase in human breast cancer cells, in comparison to non-transformed mammary epithelial cells 184A1N4 and MCF-10A. Results clearly indicated that unlike non-transformed mammary epithelial cells, breast cancer cells do not respond to the known activators of matriptase, serum and sphingosine 1-phosphate (S1P). Similar levels of activated matriptase were detected in breast cancer cells, grown in the presence or absence of S1P. However, up to five-fold higher levels of activated matriptase were detected in the conditioned media from the cancer cells grown in the absence of serum and S1P, when compared to non-transformed mammary epithelial cells. S1P also induces formation of cortical actin structures in non-transformed cells, but not in breast cancer cells. These results show that in non-transformed cells, S1P induces a rearrangement of the actin cytoskeleton and stimulates proteolytic activity on cell surfaces. In contrast, S1P treatment of breast cancer cells does not activate matriptase, and instead these cells constitutively activate the protease. In addition, breast cancer cells respond differently to S1P in terms of the regulation of actin cytoskeletal structures. Matriptase and its cognate inhibitor, HGF activator inhibitor 1 (HAI-1) colocalize on the cell periphery of breast cancer cells and form stable complexes in the extracellular milieu, suggesting that the inhibitor serves to prevent undesired proteolysis in these cells. Finally, we demonstrate that treatment of T-47D cells with epidermal growth factor (EGF), which promotes cell ruffling, stimulates increased accumulation of activated matriptase at the sites of membrane ruffling, suggesting a possible functional role at these sites.
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PMID:Deregulated activation of matriptase in breast cancer cells. 1249 94

In this study, Charlton's and Tomihisa's methods were modified to investigate the thrombolytic effect of corilagin from the Chinese herbal plant Phyllanthus urinaria L., as well as its effect on carotid artery patency status. The activity of type 1 plasminogen activator inhibitor (PAI-1) in rat plasma or platelet-released substances and tissue-type plasminogen activator (tPA) in rat plasma was assayed by use of a chromogenic substrate. The results showed that corilagin had a dose-dependent thrombolytic effect in rats. 5 mg/kg of corilagin produced a nearly similar reperfusion rate to that of 20000 U/kg of urokinase, whereas it produced a lower reocclusion rate than urokinase. Corilagin significantly inhibited PAI-1 activity in rat plasma or platelet-released substances while it elevated plasma tPA activity, in a concentration-dependent manner. Corilagin, however, had no influence on rabbit platelet aggregation. It is indicated that corilagin inhibited PAI-1 activity and increased tPA activity, and this property of corilagin is assumed to be responsible for the thrombolytic effect. Abbreviations. PO:persistent occlusion CR:cyclic reflow PP:persistent patency PAI-1:type 1 plasminogen activator inhibitor tPA:tissue-type plasminogen activator PBS:phosphate buffer solution IC (50):50 % of inhibitory concentration PRP:platelet-rich plasma ADP:adenosine diphosphate AA:arachidonic acid PAF:platelet-activating factor
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PMID:Modulation of PAI-1 and tPA activity and thrombolytic effects of corilagin. 1475 26


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